From ed1fbae4fb7b7f293406032b54bf182e0a54701a Mon Sep 17 00:00:00 2001
From: Devyani Ravi <devyaniravi2003@gmail.com>
Date: Sun, 29 Sep 2024 18:04:27 +0000
Subject: [PATCH] Mod1_D

---
 docs/.vuepress/components/IterativeCycle.vue | 11 ++---------
 1 file changed, 2 insertions(+), 9 deletions(-)

diff --git a/docs/.vuepress/components/IterativeCycle.vue b/docs/.vuepress/components/IterativeCycle.vue
index 3b7ac5a..29f487d 100644
--- a/docs/.vuepress/components/IterativeCycle.vue
+++ b/docs/.vuepress/components/IterativeCycle.vue
@@ -67,15 +67,8 @@ export default {
                     | nasB                            | 2895 bp               |
                     | nasA + Terminator               | 2730 bp               |
 
-                    <p>DNRA, typically utilised by bacteria in anaerobic conditions for the purposes of energy conservation, involves converting NO₃⁻ into NH₄⁺ in a two-step reaction via the NO₂⁻ intermediate (Herrmann & Taubert, 2022). While DNRA retains nitrogen in its bioavailable form (NH₄⁺), it does not directly incorporate it into organic compounds. Thus, both denitrification and DNRA result in the loss of available nitrogen—either as atmospheric nitrogen in the case of denitrification or as NH₄⁺ that is not assimilated into biomass in the case of DNRA.</p>
-
-                    <p>In contrast, the assimilatory NO₃⁻ pathway leads to the incorporation of nitrogen into organic compounds, such as amino acids, conserving it within the organism (Moreno-Vivián & Flores, 2007; Jiang & Jiao, 2015). These amino acids can then be used to produce single-cell proteins (SCPs). The assimilatory pathway not only retains nitrogen but also contributes to the production of microbial biomass, thus providing a more efficient means of nitrogen utilisation.</p>
-
-                    <p>The assimilatory pathway in bacteria comprises several steps. First, NO₃⁻ is captured and internalised from the extracellular environment to the intracellular space. This step is mediated by a NO₃⁻-transporter, which is most commonly an ATP-binding cassette (ABC)-type transporter located in the cytoplasmic membrane (Moreno-Vivián & Flores, 2007). The transporter consists of three subunits: a periplasmic protein that binds NO₃⁻ with high affinity (even at low extracellular concentrations of NO₃⁻), a transmembrane protein that facilitates the transport of NO₃⁻ across the membrane, and a cytoplasmic ATPase anchored to the membrane, which hydrolyses ATP to provide energy for the process (Lin & Stewart, 1997; Moreno-Vivián & Flores, 2007).</p>
-
-                    <p>Once NO₃⁻ is internalised, it is then reduced to NO₂⁻ by assimilatory nitrate reductase (Nas). This enzyme is NADH-dependent and has two subunits: a large catalytic subunit, which contains the essential active site for the reduction of NO₃⁻, and a small NADH oxidoreductase subunit, which facilitates the transfer of electrons to the active site (Lin & Stewart, 1997; Moreno-Vivián & Flores, 2007). In the following step, NO₂⁻ is further reduced to NH₄⁺ by the monomeric nitrite reductase (Nir). Afterwards, the produced NH₄⁺ is incorporated into amino acids, specifically glutamine and glutamate, through the GS-GOGAT and GDH pathways (Moreno-Vivián & Flores, 2007; van Heeswijk et al., 2013).</p>
-
-                    <p>The GS-GOGAT pathway consists of two key steps. First, the enzyme glutamine synthetase (GS) catalyses an ATP-dependent reaction that converts glutamate to glutamine by incorporating an ammonium ion. Following this, glutamate synthase (GOGAT) transfers the amide group from glutamine to 2-oxoglutarate, resulting in the production of two glutamate molecules. In contrast, the GDH pathway employs a more direct approach. The enzyme glutamate dehydrogenase (GDH) catalyses the incorporation of an ammonium ion (NH₄⁺) directly into 2-oxoglutarate, forming glutamate in a single step. The resulting amino acids, glutamate and glutamine, undergo further transamidation and transamination, yielding various amino acids, which then serve as building blocks for the biosynthesis of proteins during translation (van Heeswijk et al., 2013).</p>
+                    <p>For linearisation of the pSEVA261 plasmid, PCR primers have been designed. These primers contain sequences that are complementary to the overhangs of Fragment 1 and Fragment 4. Notably, the primers also exclude the multiple cloning site, which is in turn replaced by the fragments.</p>
+                    <p>Each fragment has a corresponding ribosome binding site (RBS) positioned immediately upstream of the start codon to ensure proper translation. The native P1 promoter is located upstream of the gene fragments to initiate transcription. The terminator is introduced downstream of the gene sequences for proper transcription termination. Since the gBlocks have been divided into four segments, an additional set of eight primers have been designed with sequences complementary to adjacent fragments, ensuring proper alignment for Gibson assembly. Each primer adds approximately 40 base pairs of overlap, following guidelines from SnapGene, to facilitate seamless assembly of the fragments. The final recombinant plasmid has a total length of 12,413 base pairs.</p> 
                     <img src="https://static.igem.wiki/teams/5306/aa-pathway.png" alt="Amino Acid Pathway" style="width:100%; height:auto;" />
                     <figcaption style="font-style: italic;">
                     Figure X. The pathways that result in the biosynthesis of glutamine and glutamate. The GDH pathway is shown in the left panel. The GS-GOGAT pathway is shown in the right panel. 
-- 
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